Expression of the gus A gene in embryogenic cell lines of Sitka spruce following Agrobacterium-mediated transformation

Transformation of Picea sitchensis (Bong) Carr. was investigated by incubating embryogenic cell lines, initiated from immature and mature zygotic embryos, with a supervirulent strain of Agrobacterium tumefaciens. The latter carried a gus A-intron gene. Transient gene expression was determined histochemically by recording the number of distinct areas of beta-glucuronidase (GUS) activity. Maximum expression of the gus gene was achieved with a bacterial suspension with an OD600 of 0.8-1.1 diluted with an equal volume of MPM medium, inoculation of cells with bacteria for 30 min, 72 h co-cultivation period and exposure of Agrobacterium and plant cells to 50 mu M acetosyringone. These results are discussed in relation to Agrobacterium-mediated gene delivery for the stable transformation of Sitka spruce and other conifers.